Methods of treating or preventing ibd with il-18

ABSTRACT

The present invention relates generally to the use of IL-18, also known as interferon-γ-inducing factor (IGIF), in the prevention and/or treatment of inflammatory bowel diseases.

FIELD OF THE INVENTION

The present invention relates generally to the use of IL-18, also knownas interferon-γ-inducing factor (IGIF), in the prevention and/ortreatment of inflammatory bowel diseases.

BACKGROUND OF THE INVENTION

IL-18 is a recently discovered novel cytokine. Active IL-18 contains 157amino acid residues. It has potent biological activities, includinginduction of interferon-γ-production by T cells and splenocytes,enhancement of the killing activity of NK cells and promotion of thedifferentiation of naive CD4⁺T cells into Th1 cells. In addition, humanIL-18 augments the production of GM-CSF and decreases the production ofIL-10. IL-18 has been shown to have greater interferon-γ inducingcapabilities than IL-12, and appears to have different receptors andutilize a distinct signal transduction pathway.

CD4⁺ T cells are the central regulatory elements of all immuneresponses. They are divided into two subsets, Th1 and Th2. Each subsetis defined by its ability to secrete different cytokines. Interestingly,the most potent inducers for the differentiation are cytokinesthemselves. The development of Th2 cells from naive precursors isinduced by IL-4. Prior to the discovery of IL-18, IL-12 was thought ofas the principal Th1 inducing cytokine. IL-18 is also a Th1 inducingcytokine and is more potent than IL-12 in stimulating the production ofinterferon-γ.

Th1 cells secrete IL-2, interferon-β, and TNF-β. Interferon-γ, thesignature Th1 cytokine, acts directly on macrophages to enhance theirmicrobiocidal and phagocytic activities. As a result, the activatedmacrophages can efficiently destroy intracellular pathogens and tumorcells. The Th2 cells produce IL-4, IL-5, IL-6, IL-10 and IL-13, whichact by helping B cells develop into antibody-producing cells. Takentogether, Th1 cells are primarily responsible for cell-mediatedimmunity, while Th2 cells are responsible for humoral immunity.

IL-18, the encoding nucleotide sequence and certain physicochemicalchemical properties of the purified protein is known.

Kabushiki Kaisha Hayashibara Seibutsu Kayaku Kenkyujo's (“Hayashibara”),U.S. Pat. No. 5,912,324, which corresponds to EP 0 692 536 published onJan. 17, 1996, discloses a mouse protein which induces IFN-gammaproduction by immunocompetent cells, the protein being furthercharacterized as having certain physicochemical properties and a definedpartial amino acid sequence. Also disclosed is a protein having a 157 aasequence, two fragments thereof, DNA (471 bp) encoding the protein,hybridomas, protein purification methods, and methods for detecting theprotein.

Hayashibara's U.S. Pat. No. 6,214,584, which corresponds to EP 0 712 931published on May 22, 1996, discloses a 157 aa human protein andhomologues thereof, DNA encoding the protein, transformants, processesfor preparing the protein, monoclonal antibodies against the protein,hybridomas, protein purification methods, methods for detecting theprotein, and methods of treatment and/or prevention of malignant tumors,viral diseases, bacterial infectious diseases, and immune diseases.

Incyte Pharmaceuticals, Inc.'s, WO 97/24441, published on Jul. 10, 1997,discloses a 193 aa protein corresponding to IL-18 precursor and encodingDNA.

Inflammatory bowel disease (IBD) is a group of chronic disorders thatcause inflammation in the small and large intestine. IBD includesCrohn's disease and ulcerative colitis. Further, IBD can also includeinflammatory colitis caused by bacteria, ischemia, radiation, drugs orchemical substances.

The present invention relates to the use of a IL-18 polypeptide for thetreatment or prevention of IBD, including, but not limited to Crohn'sdisease, ulcerative colitis, and inflammatory colitis caused bybacteria, ischemia, radiation, drugs or chemical substances.

SUMMARY OF THE INVENTION

In one aspect, the present invention provides a method of treating orpreventing IBD, including, but not limited to Crohn's disease,ulcerative colitis, and inflammatory colitis caused by bacteria,ischemia, radiation, drugs or chemical substances; comprising,administering a therapeutically effective amount of a IL-18 polypeptide.

In further aspect, the invention also relates to a pharmaceuticalcomposition comprising therapeutically effective amount of a IL-18polypeptide to treat or prevent IBD, including, but not limited toCrohn's disease, ulcerative colitis, and inflammatory colitis caused bybacteria, ischemia, radiation, drugs or chemical substances, and apharmaceutically acceptable carrier.

Yet in a further aspect, the present invention relates to the use of aIL-18 polypeptide in the preparation of a medicament for the treatmentor prevention of IBD, including, but not limited to Crohn's disease,ulcerative colitis, and inflammatory colitis caused by bacteria,ischemia, radiation, drugs or chemical substances.

DETAILED DESCRIPTION OF THE INVENTION

The following definitions are provided to facilitate understanding ofcertain terms and abbreviations used frequently in this application.

“Identity,” as known in the art, is a relationship between two or morepolypeptide sequences or two or more polynucleotide sequences, asdetermined by comparing ihe sequences. In the art, “identity” also meansthe degree of sequence relatedness between polypeptide or polynucleotidesequences, as the case may be, as determined by the match betweenstrings of such sequences. “Identity” and “similarity” can be readilycalculated by known methods, including but not limited to thosedescribed in (Computational Molecular Biology, Lesk, A. M., ed., OxfordUniversity Press, New York, 1988; Biocomputing: Informatics and GenomeProjects, Smith, D. W., ed., Academic Press, New York, 1993; ComputerAnalysis of Sequence Data, Part I, Griffin, A. M., and Griffin, H. G.,eds., Humana Press, New Jersey, 1994; Sequence Analysis in MolecularBiology, von Heinje, G., Academic Press, 1987; and Sequence AnalysisPrimer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York,1991; and Carillo, H., and Lipman, D., SIAM J. Applied Math., 48: 1073(1988). Preferred methods to determine identity are designed to give thelargest match between the sequences tested. Methods to determineidentity and similarity are codified in publicly available computerprograms. Preferred computer program methods to determine identity andsimilarity between two sequences include, but are not limited to, theGCG program package (Devereux, J., et al., Nucleic Acids Research 12(1):387 (1984)), BLASTP, BLASTN, and FASTA (Atschul, S. F. et al., J. Molec.Biol. 215: 403-410 (1990). The BLAST X program is publicly availablefrom NCBI and other sources (BLAST Manual, Altschul, S., et al., NCBINLM NIH Bethesda, Md. 20894; Altschul, S., et al., J. Mol. Biol. 215:403-410 (1990). The well known Smith Waterman algorithm may also be usedto determine identity.

“Isolated” means altered “by the hand of man” from the natural state. Ifan “isolated” composition or substance occurs in nature, it has beenchanged or removed from its original environment, or both. For example,a polynucleotide or a polypeptide naturally present in a living animalis not “isolated,” but the same polynucleotide or polypeptide separatedfrom the coexisting materials of its natural state is “isolated”, as theterm is employed herein.

“Polypeptide” refers to any peptide or protein comprising two or moreamino acids joined to each other by peptide bonds or modified peptidebonds, i.e., peptide isosteres. “Polypeptide” refers to both shortchains, commonly referred to as peptides, oligopeptides or oligomers,and to longer chains, generally referred to as proteins. Polypeptidesmay contain amino acids other than the 20 gene-encoded amino acids.“Polypeptides” include amino acid sequences modified either by naturalprocesses, such as post-translational processing, or by chemicalmodification techniques which are well known in the art. Suchmodifications are well described in basic texts and in more detailedmonographs, as well as in a voluminous research literature.Modifications may occur anywhere in a polypeptide, including the peptidebackbone, the amino acid side-chains and the amino or carboxyl termini.It will be appreciated that the same type of modification may be presentto the same or varying degrees at several sites in a given polypeptide.Also, a given polypeptide may contain many types of modifications.Polypeptides may be branched as a result of ubiquitination, and they maybe cyclic, with or without branching. Cyclic, branched and branchedcyclic polypeptides may result from post-translation natural processesor may be made by synthetic methods. Modifications include acetylation,acylation, ADP-ribosylation, amidation, covalent attachment of flavin,covalent attachment of a heme moiety, covalent attachment of anucleotide or nucleotide derivative, covalent attachment of a lipid orlipid derivative, covalent attachment of phosphotidylinositol,cross-linking, cyclization, disulfide bond formation, demethylation,formation of covalent cross-links, formation of cystine, formation ofpyroglutamate, formylation, gamma-carboxylation, glycosylation, GPIanchor formation, hydroxylation, iodination, methylation,myristoylation, oxidation, proteolytic processing, phosphorylation,prenylation, racemization, selenoylation, sulfation, transfer-RNAmediated addition of amino acids to proteins such as arginylation, andubiquitination (see, for instance, PROTEINS—STRUCTURE AND MOLECULARPROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, NewYork, 1993; Wold, F., Post-translational Protein Modifications:Perspectives and Prospects, pgs. 1-12 in POSTTRANSLATIONAL COVALENTMODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York,1983; Seifter et al., “Analysis for protein modifications and nonproteincofactors”, Meth Enzymol (1990) 182: 626-646 and Rattan et al., “ProteinSynthesis: Post-translational Modifications and Aging”, Ann NY Acad Sci(1992) 663: 48-62).

“Variant” refers to a polynucleotide or polypeptide that differs from areference polynucleotide or polypeptide, but retains essentialproperties. A typical variant of a polynucleotide differs in nucleotidesequence from another, reference polynucleotide. Changes in thenucleotide sequence of the variant may or may not alter the amino acidsequence of a polypeptide encoded by the reference polynucleotide.Nucleotide changes may result in amino acid substitutions, additions,deletions, fusions and truncations in the polypeptide encoded by thereference sequence, as discussed below. A typical variant of apolypeptide differs in amino acid sequence from another, referencepolypeptide. Generally, differences are limited so that the sequences ofthe reference polypeptide and the variant are closely similar overalland, in many regions, identical. A variant and reference polypeptide maydiffer in amino acid sequence by one or more substitutions, additions,deletions in any combination. A substituted or inserted amino acidresidue may or may not be one encoded by the genetic code. A variant ofa polynucleotide or polypeptide may be a naturally occurring such as anallelic variant, or it may be a variant that is not known to occurnaturally. Non-naturally occurring variants of polynucleotides andpolypeptides may be made by mutagenesis techniques or by directsynthesis.

Preferred parameters for polypeptide sequence comparison include thefollowing:

-   1) Algorithm: Needleman and Wunsch, J. Mol. Biol. 48: 443-453 (1970)    Comparison matrix: BLOSSUM62 from Hentikoff and Hentikoff, Proc.    Natl. Acad. Sci. USA. 89: 10915-10919 (1992)-   Gap Penalty: 12-   Gap Length Penalty: 4

A program useful with these parameters is publicly available as the“gap” program from Genetics Computer Group, Madison Wis. Theaforementioned parameters are the default parameters for peptidecomparisons (along with no penalty for end gaps).

A polypeptide sequence of the present invention may be identical to thereference sequence of SEQ ID NO:1 or SEQ ID NO:2, that is be 100%identical, or it may include up to a certain integer number of aminoacid alterations as compared to the reference sequence such that the %identity is less than 100%. Such alterations are selected from the groupconsisting of at least one amino acid deletion, substitution, includingconservative and non-conservative substitution, or insertion, andwherein said alterations may occur at the amino- or carboxy-terminalpositions of the reference polypeptide sequence or anywhere betweenthose terminal positions, interspersed either individually among theamino acids in the reference sequence or in one or more contiguousgroups within the reference sequence. The number of amino acidalterations for a given % identity is determined by multiplying thetotal number of amino acids in SEQ ID NO:1 or SEQ ID NO:2 by thenumerical percent of the respective percent identity (divided by 100)and then subtracting that product from said total number of amino acidsin SEQ ID NO:1 or SEQ ID NO:2, respectively, or:n _(a) ≦x _(a)−(x _(a) ·y)wherein n_(a) is the number of amino acid alterations, x_(a) is thetotal number of amino acids in SEQ ID NO:1 or SEQ ID NO:2, and y is, forinstance 0.70 for 70%, 0.80 for 80%, 0.85 for 85% etc., and wherein anynon-integer product of x_(a) and y is rounded down to the nearestinteger prior to subtracting it from x_(a).

“Fusion protein” refers to a protein encoded by two, often unrelated,fused genes or fragments thereof. In one example, EP-A-0 464 disclosesfusion proteins comprising various portions of constant region ofimmunoglobulin molecules together with another human protein or partthereof. In many cases, employing an immunoglobulin Fc region as a partof a fusion protein is advantageous for use in therapy and diagnosisresulting in, for example, improved pharmacokinetic properties [see,e.g., EP-A 0232 262]. On the other hand, for some uses it would bedesirable to be able to delete the Fc part after the fusion protein hasbeen expressed, detected and purified.

IL-18 Polypeptide

A IL-18 polypeptide is disclosed in EP 0692536A2, EP 0712931A2,EP0767178A1, and WO 97/2441. The polypeptides include isolatedpolypeptides comprising an amino acid sequence which has at least 70%identity, preferably at least 80% identity, more preferably at least 90%identity, yet more preferably at least 95% identity, most preferably atleast 97-99% identity, to that of SEQ ID NO:1 (human IL-18) and SEQ IDNO:2 (murine IL-18) over the entire length of SEQ ID NO:1 and SEQ IDNO:2, respectively. Such polypeptides include those comprising the aminoacid of SEQ ID NO:1 and SEQ ID NO:2, respectively.

Polypeptides of the present invention are interferon-γ-inducingpolypeptides. They play a primary role in the induction of cell-mediateimmunity, including induction of interferon-γ production by T cells andspleenocytes enhancement of the killing activity of NK cells andpromotion of the differentiation of naive CD4+ T cells into Th1 cells.These properties are hereinafter referred to as “IL-18 activity” or“IL-18 polypeptide activity” or “biological activity of IL-18”. Alsoincluded amongst these activities are antigenic and immunogenicactivities of said IL-18 polypeptides, in particular the antigenic andimmunogenic activities of the polypeptides of SEQ ID NO:1 and SEQ IDNO:2. Preferably, a polypeptide of the present invention exhibits atleast one biological activity of IL-18.

The polypeptides of the present invention may be in the form of the“mature” protein or may be a part of a larger protein such as a fusionprotein. It is often advantageous to include an additional amino acidsequence which contains secretory or leader sequences, pro-sequences,sequences which aid in purification such as multiple histidine residues,or an additional sequence for stability during recombinant production.

The present invention also includes variants of the aforementionedpolypeptides, that is polypeptides that vary from the referents byconservative amino acid substitutions, whereby a residue is substitutedby another with like characteristics. Typical such substitutions areamong Ala, Val, Leu and Ile; among Ser and Thr; among the acidicresidues Asp and Glu; among Asn and Gln; and among the basic residuesLys and Arg; or aromatic residues Phe and Tyr. Particularly preferredare variants in which several, 5-10, 1-5, 1-3, 1-2 or 1 amino acids aresubstituted, deleted, or added in any combination.

Polypeptides of the present invention can be prepared in any suitablemanner. Such polypeptides include isolated naturally occurringpolypeptides, recombinantly produced polypeptides, syntheticallyproduced polypeptides, or polypeptides produced by a combination ofthese methods. Means for preparing such polypeptides are well understoodin the art.

Recombinant polypeptides of the present invention may be prepared byprocesses well known in the art from genetically engineered host cellscomprising expression systems. Accordingly, in a further aspect, thepresent invention relates to expression systems which comprises apolynucleotide or polynucleotides encoding the polypeptides of thepresent invention, to host cells which are genetically engineered withsuch expression systems and to the production of polypeptides of theinvention by recombinant techniques. Cell-free translation systems canalso be employed to produce such proteins using RNAs derived from theDNA constructs of the present invention.

Representative examples of appropriate hosts include bacterial cells,such as streptococci, staphylococci, E. coli, Streptomyces and Bacillussubtilis cells; fungal cells, such as yeast cells and Aspergillus cells;insect cells such as Drosophila S2 and Spodoptera Sf9 cells; animalcells such as CHO, COS, HeLa, C127, 3T3, BHK, HEK 293 and Bowes melanomacells; and plant cells.

A great variety of expression systems can be used, for instance,chromosomal, episomal and virus-derived systems, e.g., vectors derivedfrom bacterial plasmids, from bacteriophage, from transposons, fromyeast episomes, from insertion elements, from yeast chromosomalelements, from viruses such as baculoviruses, papova viruses, such asSV40, vaccinia viruses, adenoviruses, fowl pox viruses, pseudorabiesviruses and retroviruses, and vectors derived from combinations thereof,such as those derived from plasmid and bacteriophage genetic elements,such as cosmids and phagemids. The expression systems may containcontrol regions that regulate as well as engender expression. Generally,any system or vector which is able to maintain, propagate or express apolynucleotide to produce a polypeptide in a host may be used. Theappropriate nucleotide sequence may be inserted into an expressionsystem by any of a variety of well-known and routine techniques, suchas, for example, those set forth in Sambrook et al., Molecular Cloning:A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory Press, ColdSpring Harbor, N.Y. (1989). Appropriate secretion signals may beincorporated into the desired polypeptide to allow secretion of thetranslated protein into the lumen of the endoplasmic reticulum, theperiplasmic space or the extracellular environment. These signals may beendogenous to the polypeptide or they may be heterologous signals.

Polypeptides of the present invention can be recovered and purified fromrecombinant cell cultures by well-known methods including ammoniumsulfate or ethanol precipitation, acid extraction, anion or cationexchange chromatography, phosphocellulose chromatography, hydrophobicinteraction chromatography, high performance liquid chromatography,hydroxylapatite chromatography and lectin chromatography. Mostpreferably, affinity chromatography is employed for purification.Well-known techniques for refolding proteins may be employed toregenerate active conformation when the polypeptide is denatured duringisolation and or purification.

The present invention also provides for pharmaceutical compositionscomprising a therapeutically effective amount of IL-18, optionally incombination with another agent. Pharmaceutically acceptable carriers orexcipients may also be employed. The pharmaceutical carrier employed maybe, for example, either a solid or a liquid. Exemplary of solid carriersinclude, but are not limited to lactose, terra alba, sucrose, talc,gelatin, agar, pectin, acacia, magnesium stearate, stearic acid and thelike. Exemplary of liquid carriers include, but are not limited to,saline, buffered saline, dextrose, water, glycerol, ethanol syrup,peanut oil olive oil, and combinations thereof. Similarly, the carrieror diluent may include time delay material well known in the art, suchas glyceryl monostearate or glyceryl distearate alone or with a waxethylcellulose, hydroxypropylmethylcellulose, methylmethacrylate and thelike.

The invention further relates to pharmaceutical packs and kitscomprising one or more containers filled with one or more of theingredients of the aforementioned compositions of the invention. Thepolypeptides may be employed alone or in conjunction with othercompounds, such as therapeutic compounds.

The composition will be adapted to the route of administration, forinstance by a systemic or an oral route. Preferred forms of systemicadministration include injection, typically by intravenous injection.Other injection routes, such as subcutaneous, intramuscular, orintraperitoneal, can be used. In addition, if the present invention canbe formulated in an enteric or an encapsulated formulation, oraladministration may be possible. Alternative means for systemicadministration include transmucosal and transdermal administration usingpenetrants such as bile salts or fusidic acids or other detergents.Administration of these combinations may also be topical and/orlocalized, in the form of salves, pastes, gels, and the like.

The dosage range of IL-18 required depends on the choice of adjuvant, ifany, the route of administration, the nature of the formulation, thenature of the subject's condition, and the judgment of the attendingpractitioner. Suitable dosages of the composition, however, for IL-18are in the range of 1 nanogram/kilogram to 1 milligram/kilogram ofsubject. Wide variations in the needed dosage, however, are to beexpected in view of the variety of compounds available and the differingefficiencies of various routes of administration. For example,transdermal administration would be expected to require higher dosagesthan administration by intravenous injection. Variations in these dosagelevels can be adjusted using standard empirical routines foroptimization, as is well understood in the art.

The schedule for the administration of the composition depends on thedosage, on the choice of adjuvant, the route of administration, thenature of the formulation, the nature of the subject's condition, andthe judgment of the attending practitioner. Suitable schedules foradministration, are daily, weekly, or monthly. Wide variations in theschedules for the administration of the composition, however, are to beexpected in view of the variety of other agents available and thediffering efficiencies of various routes of administration. For example,transdermal administration would be expected to require higher dosagesthan administration by intravenous injection. Variations in theseschedules for the administration of the composition can be adjustedusing standard empirical routines for optimization, as is wellunderstood in the art.

All publications, including but not limited to patents and patentapplications, cited in this specification are herein incorporated byreference as if each individual publication were specifically andindividually indicated to be incorporated by reference herein as thoughfully set forth.

It is believed that one skilled in the art can, using the precedingdescription, utilize the present invention to its fullest extent.Therefore the Examples herein are to be construed as merely illustrativeand not a limitation of the scope of the present invention in any way.

EXAMPLES Method

Induction of Colitis

Female, 8-week-old BALB/c mice (Charles River Japan) were used in thisstudy. Colitis was induced by providing drinking water containing 3%dextran sulfate sodium (DSS, ICN Biomedicals Inc., M.W.=36,000-50,000)for 5 days. The administration of DSS was discontinued on day 5, andmice were given tap water alone for 7 days until on day 12.

Evaluation of Colitis

The disease activity index (DAI) was determined in all animals, byscoring body weight, stool consistency and rectal bleeding as describedby Murthy, S. N. S. (Digestive Diseases and Sciences, 38(9) p.1722-1734(1993)). The method of scoring is shown in Table 1. Severity ofcolitis was evaluated by area under the curve (AUC) calculated based onDAI curve ranged from day 3 to day 7 (AUC (3-7 day)), from day 7 to day10 (AUC (7-10 day)), from day 10 to day 12 (AUC (10-12 day)) and fromday 0 to day 12 (AUC (0-12 day)). TABLE 1 Criteria for scoring Occultblood or Score Weight loss (%) Stool consistency gross bleeding 0 NoneNormal Negative 1 1-5 Loose stool Negative 2  5-10 Severe loose stoolHemoccult positive 3 10-15 Diarrhea Hemoccult strong positive 4 >15Severe diarrhea Gross bleedingDAI = (combined score of weight loss, stool consistency and bleeding)/3.

Experimental Design

Twelve mice were used in each group. IL-18 (SEQ ID NO: 2) was dissolvedin buffer (25 mM Na-acetate, 100 mM NaCl, 0.1 mM EDTA, 6.0%(w/v)sucrose, pH 5.5). IL-18 at 0.3 ug/head or buffer was administeredintraperitoneally once a day for 12 days from day 0.

The experimental groups were set up as follows:

-   Control*-   3% DSS+buffer-   3% DSS+IL-18 (0.3 ug/head)-   * Mice which received tap water without DSS.

Results

The effect of IL-18 on DSS-Induced Colitis

The effect of IL-18 on DSS-colitis was shown in Table 2. IL-18 (0.3ug/head, i.p. q.d.) suppressed the severity of DSS-induced colitis asexpressed by a significantly lower AUC (7-10 day), AUC (10-12 day) andAUC (0-12 day) compared with buffer-treated DSS-fed mice. TABLE 2Evaluation of colitis by AUC Groups n AUC (3-7 day) AUC (7-10 day) AUC(10-12 day) AUC (0-12 day) Control 12 0.25 +/− 0.13 0.33 +/− 0.15 0.13+/− 0.06  0.79 +/− 0.33 3% DSS + buffer 11 5.36 +/− 0.53 7.38 +/− 0.544.32 +/− 0.52 18.21 +/− 1.52 3% DSS + IL-18 12 4.61 +/− 0.41 5.25 +/−0.63** 2.25 +/− 0.52** 12.99 +/− 1.42* Inhibition (%) (14.0) (28.8)(47.9) (28.7)The data were represented as mean +/− SE.n = 11 to 12.

TABLE 3 Sequence ID NO:1 Tyr Phe Gly Lys Leu Glu Ser Lys Leu Ser Val IleArg Asn Leu Asn  1               5                  10                  15 Asp Gln ValLeu Phe Ile Asp Gln Gly Asn Arg Pro Leu Phe Glu Asp             20                  25                  30 Met Thr Asp SerAsp Cys Arg Asp Asn Ala Pro Arg Thr Ile Phe Ile         35                  40                  45 Ile Ser Met Tyr LysAsp Ser Gln Pro Arg Gly Met Ala Val Thr Ile     50                  55                  60 Ser Val Lys Cys Glu LysIle Ser Thr Leu Ser Cys Glu Asn Lys Ile65                  70                  75                  80 Ile SerPhe Lys Glu Met Asn Pro Pro Asp Asn Ile Lys Asp Thr Lys                 85                  90                  95 Ser Asp IleIle Phe Phe Gln Arg Ser Val Pro Gly His Asp Asn Lys            100                 105                 110 Met Gln Phe GluSer Ser Ser Tyr Glu Gly Tyr Phe Leu Ala Cys Glu        115                 120                 125 Lys Glu Arg Asp LeuPhe Lys Leu Ile Leu Lys Lys Glu Asp Glu Leu    130                 135                 140 Gly Asp Arg Ser Ile MetPhe Thr Val Gln Asn Glu Asp 145                 150                 155

TABLE 4 Sequence ID NO:2 Asn Phe Gly Arg Leu His Cys Thr Thr Ala Val IleArg Asn Ile Asn  1               5                  10                  15 Asp Gln ValLeu Phe Val Asp Lys Arg Gln Pro Val Phe Glu Asp Met             20                  25                  30 Thr Asp Ile AspGln Ser Ala Ser Glu Pro Gln Thr Arg Leu Ile Ile         35                  40                  45 Tyr Met Tyr Lys AspSer Glu Val Arg Gly Leu Ala Val Thr Leu Ser     50                  55                  60 Val Lys Asp Ser Lys MetSer Thr Leu Ser Cys Lys Asn Lys Ile Ile 65                  70                  75                  80 Ser PheGlu Glu Met Asp Pro Pro Glu Asn Ile Asp Asp Ile Gln Ser                 85                  90                  95 Asp Leu IlePhe Phe Gln Lys Arg Val Pro Gly His Asn Lys Met Glu            100                 105                 110 Phe Glu Ser SerLeu Tyr Glu Gly His Phe Leu Ala Cys Gln Lys Glu        115                 120                 125 Asp Asp Ala Phe LysLeu Ile Leu Lys Lys Lys Asp Glu Asn Gly Asp    130                 135                 140 Lys Ser Val Met Phe ThrLeu Thr Asn Leu His Gln Ser 145                 150                 155

1. A method of treating or preventing IBD; comprising, administering atherapeutically effective amount of a polypeptide having at least 90%identity to the amino acid sequence of SEQ ID NO:1 over the entirelength of SEQ ID NO:1.
 2. A method of treating or preventing IBD;comprising, administering a therapeutically effective amount of apolypeptide having at least 90% identity to the amino acid sequence ofSEQ ID NO:2 over the entire length of SEQ ID NO:2.
 3. A method oftreating or preventing IBD; comprising, administering a therapeuticallyeffective amount of a polypeptide having the amino acid sequence of SEQID NO:1.
 4. A method of treating or preventing IBD; comprising,administering a therapeutically effective amount of a polypeptide havingthe amino acid sequence of SEQ ID NO:2.
 5. A method of claim 1, 2, 3 or4 in which IBD is selected from the group consisting of Crohn's disease,ulcerative colitis, and inflammatory colitis caused by bacteria,ischemia, radiation, drugs or chemical substances.
 6. A pharmaceuticalcomposition for treating or preventing IBD comprising therapeuticallyeffective amount of a polypeptide having at least 90% identity to theamino acid sequence of SEQ ID NO:1 over the entire length of SEQ IDNO:1.
 7. A pharmaceutical composition for treating or preventing IBDcomprising therapeutically effective amount of a polypeptide having atleast 90% identity to the amino acid sequence of SEQ ID NO:2 over theentire length of SEQ ID NO:2.
 8. A pharmaceutical composition fortreating or preventing IBD comprising therapeutically effective amountof a polypeptide having the amino acid sequence of SEQ ID NO:1.
 9. Apharmaceutical composition of claims 6-8 in which IBD is selected fromthe group consisting of Crohn's disease, ulcerative colitis, andinflammatory colitis caused by bacteria, ischemia, radiation, drugs orchemical substances.